We propose to continue our studies of the superantigen (Sag) exotoxins produced by pathogenetic strains of Staphylococcus aureus and Streptococcus pyogenes. We will focus primarily on four members of this family, the staphylococcal enterotoxins SEA, SEB and SEE and the related streptococcal superantigen SSA, which we isolated and characterized during the initial period of support. These Sag are chosen for study because of functional and structural properties that make them particularly suitable for answering the questions posed and because of the valuable library of molecular variants of the relevant genes and their products that we have developed. Our overall objective is to enhance understanding of the interaction between bacterial parasites and mammalian hosts and the mechanisms whereby Sag alter this relationship. Five specific aims are proposed: 1) Definition of those elements of the staphylococcal enterotoxins SEA and SEE and of T cell receptors that determine their interactions with one another; 2) Characterization of class II-independent Sag presentation, including definition of the responsible cell surface ligand; 3) Comparative studies of SEB with SSA and its alleles and mutants, based on structural resolution and functional analysis in order to explore the relationship between Sag potency, MHC class II and TCR interactions; 4) Analysis of the mechanism of interaction between SEA and the affinity ligand red dye A to determine whether definition of the biochemical specificity of this interaction can provide insight into biological activities of bacterial Sag and to extend use of this strategy for the identification of additional novel Sag, e.g., from M. tuberculosis; and 5) Use of SEA insertion and deletion mutants of S. aureus to assess the role of SEA in S. aureus pathogenesis in mice and to determine effects of infection with an enterotoxin-producing strain on host immune responses. We believe that the experiments proposed are conceptually novel and the experiments designed to answer them general decisive. They can be expected to improve understanding of fundamental aspects of mammalian immune response to Sag and Sag-producing bacterial pathogens and to provide insight into pathogenetic mechanisms of Sag-mediated human disease.